The microbial conversion of phytosterols (most frequently derived from soy bean oil) by various strains of bacteria is a well known process that has been used for the commercial production of androstenedione (AD) and androstadienedione (ADD) since the mid 1970's. In general, the known fermentation process involves the propagation of a mutant of Mycobacterium in an appropriate nutrient medium, transfer of the culture to a bioreactor containing the phytosterols, and then allowing the biotranformation to AD and/or ADD over a period of approximately 120 hours. Harvesting of the fermentation broth, extraction of the latter with an organic solvent, and subsequent crystallization in an organic solvent, generally provides the products AD and/or ADD as white crystalline powders. Pertinent references that discuss the known process and summarize the earlier studies are as follows:
S. Kraychy, and R. D. Muir, U.S. Pat. No. 3,684,657 (1972). W. J. Marsheck, S. Kraychy and R. D. Muir, Appl. Microbiol., 23, 72 (1972).
A. H. Conner, M. Nagaoka, J. W. Rowe and D. Perlman, Appl. and Environ. Microbiol., 32, 310 (1976).
K. Kieslich, J. Basic Microbiol., 25, 461 (1985).
One of the problems associated with the known phytosterol bio-conversion process (which problem plagues the entire steroid industry) involves the poor solubility of the substrate, in this case a phytosterol composition, in the aqueous nutrient medium. Inadequate solubility dictates the presence of only relatively low concentrations of substrate in the nutrient medium, resulting in poor contact with the micro-organism and generally leading to low yields of end products. Long fermentation times are also typically needed to achieve any satisfactory degree of bio-conversion.
Another problem associated with the known phytosterol bio-conversion process is that the end-product of the bio-conversion from phytosterols (or compositions of phytosterols) typically contains significant amounts of both AD and ADD. Given the similar chemical structure of AD and ADD, it is difficult and expensive to subsequently separate these two steroid products from one another.
A further problem associated with the known phytosterol bio-conversion process is that the micro-organism used to effect the bio-conversion (typically a mutant of Mycobacterium) is grown and propagated in nutrient media that are typically expensive to manufacture.